For example, the identified number of miRNAs in Arabidopsis, rice, maize and wheat were 232, 491, 170 and 58, respectively . But most species-specific miRNAs are still unidentified and much fewer miRNAs from tobacco have been identified. In present study, our Solexa high-throughput sequencing of tobacco roots small RNAs revealed a diverse and complex small RNA population, and expression of the 136 conserved miRNAs and 126 new miRNAs were determined. Therefore, the miRNAs sequenced in this study can definitely provide the information of tobacco miRNAs for further study on their gene regulation function in response to topping. To assess and define a putative function for a miRNA in plant, a further step of target identification is necessary. Currently, the most efficient tool available for this is the bioinformatics approach facilitated by the high degree of homology between miRNA and its target sequences in plants . In this study, putative targets were identified for 133 out of 136 conserved miRNAs and 126 new miRNAs. Although targets can be predicted for many new miRNAs, the rate of false positives is usually higher for new miRNAs than for conserved miRNAs . Therefore, the targets of new miRNAs need to be further validated. Of these miRNAs whose targets had been identified, the miRNAs which change markedly belong to 53 families and their targets have BEZ235 PI3K inhibitor different biological functions including plant R428 development, response to stress, response to hormone, N metabolism, C metabolism, signal transduction, nucleic acid metabolism and other metabolism. 15 miRNA families may be involved in tobacco development, which is consistent with tobacco transition from reproductive to vegetative phase induced by topping. Of these families, ntamiR156f was only expressed in tobacco roots before topping. Squamosa promoter-binding protein-like is the target of ntamiR156 family. SPL genes encode plant-specific transcription factors that play important roles in plant phase transition, plant architecture and gibberellins signaling . Autophagocytosisassociated family protein is the target of nta-miRn60a, and it reduces shoot anthocyanin accumulation in response to cytokinin feeding to the roots, having a role in cytokinin regulated root-shoot communication. Leaf senescence can also be accelerated by the disruption of an Arabidopsis autophagy gene . In the study, the expression of nta-miR156 family and nta-miRn60a were significantly repressed by topping. Scarecrow is a member of the plant-specific GRAS family and plays a significant role in the radial patterning of both roots and shoots . Nta-miR171d targets the scarecrow transcription factor family protein. In the study, nta-miR171d was markedly induced by topping. Therefore topping can affect the radial patterning of tobacco roots. Since nta-miRn60a and nta-miR171d are involved in root development, it is easy to understand the increase in the activity,